A practical guide to in situ and ex situ characterisation of arabinogalactan proteins (AGPs) in fruits

BACKGROUND: Arabinogalactan proteins (AGPs) are plant cell components found in the extracellular matrix that play crucial roles in fruit growth and development. AGPs demonstrate structural diversity due to the presence of a protein domain and an expanded carbohydrate moiety. Considering their molecular structure, the modification of glycosylation is a primary factor contributing to the functional variety of AGPs. MAIN BODY: Immunocytochemical methods are used for qualitative and quantitative analyses of AGPs in fruit tissues. These include in situ techniques such as immunofluorescence and immunogold labelling for visualising AGP distribution at different cellular levels and ex situ methods such as Western blotting and enzyme-linked immunoenzymatic assays (ELISA) for molecular characterisation and quantitative detection of isolated AGPs. The presented techniques were modified by considering the structure of AGPs and the changes that occur in fruit tissues during the development and ripening processes. These methods are based on antibodies that recognise carbohydrate chains, which are the only commercially available highly AGP-specific tools. These probes recognise AGP epitopes and identify structural modifications and changes in spatio-temporal distribution, shedding light on their functions in fruit. CONCLUSION: This paper provides a concise overview of AGP research methods, emphasising their use in fruit tissue analysis and demonstrating the accessibility gaps in other tools used in such research (e.g. antibodies against protein moieties). It underscores fruit tissue as a valuable source of AGPs and emphasises the potential for future research to understand of AGP synthesis, degradation, and their roles in various physiological processes. Moreover, the application of advanced probes for AGP visualisation is a milestone in obtaining more detailed insights into the localisation and function of these proteins within fruit.